HER2/neu Th Peptide Vaccine Administered Concurrently With Trastuzumab Was Well Tolerated and Did Not Result in Additional Cardiac Toxicity
details 573 adverse events. The majority of toxicities were grade 1 or 2 (99%). There were four grade 3 toxicities, three possibly related to the treatment: injection-site reaction, fainting, and ulceration. There was one nonrelated grade 4 event, a stroke. The median LVEF before treatment was 61% (range, 46% to 72%) and post-treatment was 61% (range, 45% to 66%). Three patients (15%) had a decrease in LVEF to less than normal values on study. None developed symptoms of left ventricular dysfunction. Cardiac toxicities were grade 1 and 2.
HER2/neu Th Peptide Vaccine Administered Concurrently With Trastuzumab Stimulates or Boosts HER2/neu-Specific Immunity in the Majority of Patients
The median peptide-specific T-cell response before the first vaccine was a frequency of less than 1 antigen-specific cell in 75,000 PBMCs (range, 1:400,000 to 1:77,000; A). Ninety percent of patients developed new or augmented immunity. The maximal response to p369.15 was a median frequency of one in 4,121 PBMCs (range, 1:323 to 1:2,000,000; P = .0015 compared with prevaccination), the maximal response to p688.15 was one in 4,152 PBMCs (range, 1:307 to 1:3,000,000; P = .0012), and the maximal response to p971.15 was one in 2,086 PBMCs (range, 1:266 to 1:86,960; P = .0066; A). Ten (53%) of 19 patients had pre-existing immunity to any of these peptides. Sixteen patients (84%) significantly augmented immunity, three (16%) did not augment, and none had a decrease in peptide-specific immunity with immunization. The percentage of responding patients for the peptides is shown (B).
Fig 1. A human epidermal growth factor receptor 2 (HER2)/neu T-helper peptide vaccine administered concurrently with trastuzumab stimulates or boosts HER2/neu-specific immunity in the majority of patients. (A) Prevaccine (Pre) and maximal (Max) responses 1/frequency (more ...)
Peptides were derived from both the HER2/neu ECD and ICD. Of note, seven (64%) of 11 patients had significant pre-existing immunity to the ECD (median, 1:17,729; range, 1:660 to 1:3,000,000), and six (32%) of 19 patients had significant pre-existing immunity to the ICD (median, 1:309,524; range, 1:226 to 1:3,000,000; A). Sixty-nine percent of patients developed new or augmented immunity: 37% to ECD, 53% to ICD, and 21% to both. The maximal response to the ECD was a median of one cell in 2,312 PBMCs (range, 1:678 to 1:1,000,000; P = .3017 compared with baseline) and to the ICD was one cell in 9,677 PBMCs (range, 1:232 to 1:3,000,000; P = .0894; A). Five patients did not augment, and none had a significant decrease in domain immunity with immunization. The percentage of responding patients is shown (B).
Vaccination Can Elicit Tumor-Specific Cytotoxic T Cells
Embedded within the native sequence of the Th peptides are HLA-A2 binding motifs: p369.9, p688.9, and p972.9.1,5
The maximal response to class I peptides was a median frequency to p369.9 of 1:10,200 (range, 1:1,032 to 1:3,000,000; P
= .0030 compared with baseline), to p689.9 was 1:10,050 (range, 1:1,039 to 1:3,000,000; P
= .1720), and to p971.9 was 1:5,659 (range, 1:845 to 1:3,000,000; P
= .0126; A). T-cell cultures were established for five patients who had excess PBMCs available. The resultant T-cell lines could specifically lyse HER2/neu
-positive/HLA-A2–positive breast cancer cells (range, 7% to 70% lysis; B).
Fig 2. Vaccination can elicit tumor-specific cytotoxic T cells. (A) Prevaccine (Pre) and maximal (Max) responses 1/frequency (Y axis) HLA-A2 peptides (X axis). Connected points: mean and SE of six replicates with median bar. (B) Percent lysis: SKBR3 (blue), (more ...)
Seven (37%) of 19 patients had pre-existing immunity to these HLA-A2 peptides. Overall, 14 patients (74%) significantly augmented the class I HER2/neu peptide-specific immune response, four patients (21%) did not augment, and one patient had a decrease in immunity to the peptides with immunization. Percentage of responding patients for the HLA-A2 peptides is shown (C).
Vaccination-Induced or Enhanced Epitope Spreading Was Observed in the Majority of Patients and Was Associated With a Decrease in Serum TGF-ß
p98.15 and p776.15 are native epitopes of HER2/neu
, immunogenic, and not included in the vaccine formulation.11
Development of immunity to these epitopes demonstrates intramolecular epitope spreading, which was elicited or augmented in the majority of patients (A).6
The median maximal T-cell response to nonimmunizing epitopes was a frequency to p98.15 of one in 7,558 PBMCs (range, 1:1,205 to 1:3,000,000; P
= .0055 compared with prevaccination) and to p776.15 of one in 2,183 PBMCs (range, 1:527 to 1:40,000; P
= .0006; A). Nine (47%) of 19 patients had pre-existing immunity to these peptides. Fourteen patients (74%) significantly augmented the immune response, five patients (26%) did not augment, and none significantly decreased immunity to these peptides with immunization.
Fig 3. Vaccination-induced epitope spreading occurred in the majority of patients and was associated with a decrease in serum transforming growth factor beta (TGF-β) levels. (A) Prevaccine (Pre) and maximal (Max) intramolecular epitope spreading (IMS) (more ...)
We have identified several immunogenic breast cancer–associated proteins and questioned whether new or augmented immunity to IGFBP-2, p53, and topoisomerase II-α were stimulated with vaccination (ie, intermolecular epitope spreading).10,12
Seven patients had sufficient PBMCs available for this analysis. All patients had a pre-existing immune response to at least one antigen, and all seven developed new or augmented immunity to at least one of the antigens (B). The postvaccination median response to IGFBP-2 was a frequency of one in 5,405 PBMCs (range, 1:1,993 to 1:150,000; P
= .0973 compared with prevaccination), postvaccination median response to p53 was one in 6,793 PBMCs (range, 1:1,061 to 1:109,091; P
= .1282), and postvaccination median response to topoisomerase II-α was one in 3,659 PBMCs (range, 1:1,575-1:8,219; P
= .0111; B). Five patients (71%) augmented immunity to IGFBP-2, six patients (86%) augmented immunity to p53, and all patients tested augmented immunity to topoisomerase II-α. Two patients had a significant decrease in a pre-existing immune response to IGFBP-2 with immunization.
The multiple specificities of IFN-γ secreting T cells induced by vaccination led us to question whether these T cells, which could potentially traffic to tumor, might impact the immunosuppressive environment that has been described in breast cancer.13,14
TGF-ß has been shown to be elevated in the serum of patients with breast cancer and is associated with T-cell dysfunction.15–17
The greater the magnitude of the intramolecular epitope spreading T-cell response, the greater the decrease in serum TGF-ß (r
= 0.614; P
= .0003; C). There was weak correlation between the magnitude of the tetanus toxoid response, evaluated as a control, and change in TGF-ß levels (r
= 0.016; P
= .93; D).
Treg levels were measured before and after immunization in eight patients. The median percent Treg was 1.64% before immunization (range, 0.33% to 7.33%) and 1.32% 1 month after vaccines (range, 0.41% to 5.08%; P = .60). At 1 year after immunization, the median Treg level was 1.11% (range, 0.24% to 4.91%; P = .61 compared with preimmunization).
HER2/neu-Specific Immunity Can Persist and Even Increase After Active Immunizations Have Ended
shows the OS and PFS of the study population from the time of first vaccination. The median follow-up among survivors was 36 months (range, 21 to 49 months). The median PFS was 17.7 months, and the Kaplan-Meier estimate of PFS was 33% at 3 years. The median OS has not been reached; the Kaplan-Meier estimate of OS is 86% at 4 years.
Overall survival (OS) and progression-free survival (PFS) of immunized patients. Kaplan-Meier curves of the percent OS and PFS in months from the time of first vaccine (n = 21).
Although the generation of a new or augmentation of a pre-existing immune response to either immunizing peptides or protein (P = .11) or to epitope spreading peptides (P = .47) was not associated with survival, the magnitude of immunity generated tended to be higher in surviving patients. All 10 patients who had T-cell responses greater than the median to HER2/neu immunizing peptides and associated protein (P = .08) or to peptides associated with intramolecular epitope spreading (P = .09) were survivors as compared with patients with responses less than the median.
We monitored immunity to HER2/neu-related antigens after the end of immunizations in 11 patients (ECD, n = 6). Five patients (46%) maintained the same level of immunity to p369.15 in long-term follow-up as compared with the maximal response achieved during active immunization. Two (18%) decreased immunity and four (36%) continued to augment immunity (median, 1:4,121; range, 1:788 to 1:3,000,000). Seven patients (64%) maintained the same level of immunity to p688.15. One patient (9%) decreased compared with the maximal response achieved, and three patients (27%) continued to augment immunity to p688.15 (median, 1:12,876; range, 1:307 to 1:3,000,000). Five patients (46%) maintained the same level of immunity to p971.15. Two patients (18%) decreased and four patients (36%) continued to augment immunity (median, 1:5,236; range, 1:266 to 1:222,222).
Persistent immunity to the HER2/neu ECD and ICD was assessed. Five (83%) of six patients maintained the same level of immunity to the ECD in follow-up. One patient (17%) had decreased and none continued to augment immunity to the ECD (median, 1:4,625; range, 1:803 to 1:11,321). Six (55%) of 11 patients maintained immunity to the ICD. Two (18%) decreased and three (27%) continued to augment immunity (median, 1:6,024; range, 1:232 to 1:3,000,000). Finally, intramolecular epitope spreading was maintained in five patients (46%) as evidenced by immunity to p776.15. Two patients (18%) significantly decreased and four (36%) continued to augment immunity (median, 1:2,667; range, 1:527 to 1:300,000).